The objective of this proposal is to determine the three-dimensional structure of a DNA-binding protein at 5 A resolution by electron microscopy. Gp32 I, with a molecular weight of 27,000 obtained by partial proteolysis of the gene 32 product (gp32) of T4 bacteriophage, will be used for the structural investigation. Gp32 I has been found to be a stronger DNA helix destabilizer than gp32. Recent success of preparing gp32 I on a thin and two-dimensionally large crystal has made it a suitable specimen for structural determination by high resolution electron microscopy. The effect of specimen damage due to irradiating electrons will be minimized by using low dose recording techniques, and by using an "optimal" emulsion for recording. The crystallinity of the specimen would be preserved in the microscope by embedding in glucose or glucose-uranyl acetate. Because of the relatively thin specimen (less than 100 A), a weak phase approximation will be used for data interpretation. The projected potential function of the protein crystal will be calculated from the electron diffraction intensities and the image intensities. By combining the data obtained at various specimen tilting angles, a three-dimensional structure will then be reconstructed. It is expected to determine the molecular features of the secondary and tertiary conformation of gp32 I at 5 A resolution, which would provide useful information about the molecular mechanism of its function. In addition, an experimental attempt will be made to co-crystallize the gp32 I with an oligonucleotide. The structural data from gp32 I-oligonucleotide crystal would provide further information about the molecular interaction between the nucleic acid and protein, as well as the possible conformational change of gp32 I resulted from the nucleic acid binding.